This application is for revision of the study aims of R01 NR 014479-02 Microbiome and Pain in IBS (Parent Grant) to include a focus on the additional measure of fecal metabolome. Irritable bowel syndrome (IBS) affects 10-20% of adults worldwide exerting a tremendous economic, social, and emotional burden. We are studying biomarkers (gut microbiota and permeability, serum immune markers, and targeted genetic polymorphisms [Parent Grant] and in this revised aim fecal tryptophan (TRY) metabolites and bile acids (BA) that likely characterize subsets of adults with IBS. This revised aim is innovative because we will add metabolomics analyses to other clinical and microbiota analyses to characterize pathobiologically what heretofore has been primarily a phenotypically-defined condition. Building on prior serum studies we will conduct targeted fecal metabolomics analyses focused on those TRY metabolites. We are currently conducting the following 3 aims of the Parent Study. Aim 1. Compare stool GI microbiota composition, GI permeability, and pro- and antiinflammatory cytokines (GI biomarkers) in adults (18-45 yr of age) with IBS (n=100) versus (vs) Healthy Controls (HC) (n=50) without IBS. Aim 2. Among individuals with IBS, compare abdominal pain, other IBS symptoms and psychological distress symptoms in those with abnormal vs. normal GI biomarkers. The HC group results will be used to determine normal ranges for permeability, cytokines, and cluster analyses will be used to identify IBS subtypes based on type of microbiota community. Aim 3. Explore patterns of associations among GI biomarkers (permeability, cytokines, microbiota); and associations of GI biomarkers with abdominal pain, other IBS symptoms, stress, psychological distress, genetic polymorphisms (SERT, IL-10, IL-10R). The goal is to identify IBS subgroups based on combinations of biomarkers and clinical phenotypes, which could potentially correspond to different etiologies. Cluster analyses will be applied to microbiota data to identify subtypes with different microbiota profiles. Revised Aim 4: Explore fecal TRP metabolites and BAs, preliminarily compare IBS and HCs and examine patterns of association similar to that described in Aim 3 for microbiome. Aims 3 & 4 are exploratory and any proposed subtypes will need validation in an independent sample. Our proposal addresses NINR goals including developing new diagnostic tools and predictive biomarkers, encouraging risk taking, innovation, and creativity, to address a healthcare cost (costs are 57% greater for IBS). The proposed revision is important because providers are challenged given the pathobiology of IBS is poorly defined and treatments are not universally effective. Future studies could target management with abnormal biomarkers differently from those with no biomarker abnormalities but greater psychological distress.